DESCRIPTION (Applicant's abstract): A novel isothermal DNA amplification technique has been developed called cascade rolling circle amplification (CRCA). This method is simple, rapid, and highly sensitive, achieving over a billion-fold amplification within one hour. By making one of the primers an energy transfer-labeled primer that generates a fluorescent signal only when incorporated into the amplicon, CRCA products can be detected within a closed system. These features make this technology suitable for a variety of diagnostic applications including both automated platforms and point-of-care field devices. Phase I research has demonstrated the feasibility of CRCA for DNA diagnostics, including detection of pathogenic agents. The objectives of this Phase II project include i) improvements in the assay to make the technology more adaptable to a variety of platforms, ii) development of multiplex assay using energy transfer-labeled primers of different colors, iii) optimizing the system for real-time detection to accurately quantitate target sequences, iv) adapting CRCA to a point-of-care diagnostic platform that is non-fluorescence based, and v) adapting the technology to an array platform. Long-range objectives include implementing CRCA in an automated amplification/detection system and developing broad applications for the point-of-care diagnostic market PROPOSED COMMERCIAL APPLICATION: NOT AVAILABLE